Abstract The objectives of this project are first, to increase the efficacy and safety of live poxvirus vaccines against smallpox and other virulent poxviruses, and second, to increase the efficacy of live vaccinia virus vaccines as a vaccine platform for protection against multiple agents important in the fields of biodefense and emerging infections. The specific aims of this project are as follows: 1. To identify key mechanisms of viral induction of immune responses, particularly those contributing to the efficacy of vaccinia virus MVA vaccines: The goal is to learn how to maximize the induction of protective responses through the manipulation of viral control of these processes. 2. To determine if different orthopoxviruses, specifically vaccinia virus MVA, vaccinia virus (WR), and cowpox virus differentially affect dendritic cell (DC) responses to viral infection. The goal is to determine the mechanisms contributing to species-specific effects upon DC functions. We will use this information to manipulate viral control of DC responses to infection in order to maximize protective responses induced by vaccine viruses. 3. To determine if the efficacies of MVA vaccines can be improved by modification of accessory proteins encoded by the vaccine virus. The goal is to determine if the stability, delivery regimen, and the efficacy of vaccinia virus MVA vaccines can be improved by the expression of two cowpox/vaccinia virus accessory genes that encode proteins enabling the MVA virus vaccine to embed its intracellular mature virus (IMV) within A-type inclusion particles. The current MVA virus vaccine provides a safe alternative to the currently licensed smallpox vaccines that are associated with a significant degree of adverse effects. However, because of the MVA virus is defective for replication in human cells, large doses of MVA are needed, and multiple doses may be needed for effective protection against specific agents. Therefore, methods to enhance the stability and delivery of this vaccine, as well as methods to enhance the immunogenicity of this vaccine virus, preferably to the point of reducing the vaccination to a single dose, would significantly enhance the efficacy of this vaccine vector.